Thromboxane A2(TXA2) and its precursor prostaglandin H2(PGH2), which are synthesized by platelets, are potent stimulators of platelet aggregation and constrictors of vascular smooth muscle. Thromboxane A2 has been implicated as an important pathogenetic humoral agent in a variety of cardiovascular diseases. Recently, we and others have identified a putative TXa2/PGH2 receptor in human platelets using radioligand binding assays. We have developed a radioligand binding assay for the putative human platelet TXA2/PGH2 receptor using the TXA2/PGH2 receptor antagonist [125I]-PTA-OH, (16-(3-iodo-4-hydroxyphenyl)-13,14-dihydro-15AlphaBeta-hydroxy-13-aza-Omega- tetranor-pinane-TXA2 which was synthesized in our laboratories. This proposal has three major objectives. The first is to characterize the binding of agonists and antagonists and factors which modulate their binding to the human platelet TXA2/PGH2 receptor. As a component of this objective the subcellular localization of the receptor will be determined using 3 different but complimentary approaches. These are; 1) radioligand binding of [125I]-PTA-OH to subcellular fractions of the platelet; 2) binding of a photolyzable [125I]-ligand to subcellular fractions and 3) immunocytochemical localization of antibodies directed at the receptor. The second major objective is to isolate and purify to homogeneity the human platelet TXA2/PGH2 receptor. The purified receptor will be used to 1) determine its second messenger through reconstitution in liposomes and selected cell lines and 2) determine its structural characteristics and relation to function using a variety of biophysical, biochemical and immunologic approaches. The immunologic approach to be carried out through the use of a bank of monoclonal antibodies prepared against the receptor. We will also synthesize new TXA2/PGH2 agonists which will be radiolabelled with 125I and used for characterization of the receptor in radioligand binding assays. Finally, the receptor will be characterized in patients with altered platelet responses to arachidonic acid metabolites and their stable mimetics. A more comprehensive understanding of the structure and function of the human platelet TXA2/PGH2 receptor may lead to the improved design of drugs specifically targeted against this receptor.